Chitosan (poly-1,4-β-D-glucosamine) is a non-toxic (Journal of Biomedical Material Research 59, 2002, 585) and biodegradable (Biomaterials 20, 1999, 175) polysaccharide that is derived from chitin by deacetylation in basic conditions. The term chitosan is used to describe a wide variety of heteropolymers of glucosamine and N-acetylglucosamine with varying degrees of deacetylation and molecular weights. Chitosan has many potential applications in various fields, e.g., in pharmacy and medicine (Drug Development and Industrial Pharmacy 24, 1998, 979; Pharmaceutical Research 15, 1998, 1326; S.T.P. Pharma Science 10, 2000, 5), food science (International Dairy Journal 14, 2004, 273; Agro Food Industry Hi-Tech 14, 2003, 39), water purification (Water Research 34, 2000, 1503), pulp and paper industry (Journal of Applied Polymer Science 91, 2004, 2642), and in the textile industry (Journal of Macromolecular Science Polymer Reviews C43, 2003, 223).
The main obstacle to the use of chitosan in different applications is poor solubility properties, especially the poor aqueous solubility. The poor solubility of chitin and chitosan is due to strong intra- and intermolecular hydrogen bonding leading to highly crystallized structures. Chitosan dissolves only in acidic aqueous solutions due to the protonation of the amino groups in the polymer. Chitosan is poorly soluble in all common organic solvents. Chitosan becomes soluble in water when the degree of deacetylation is approximately 50% due to unfavourable conformation to form intermolecular hydrogen bonds (Biomacromolecules 1, 2000, 609). Various chitin and chitosan derivatives have been designed and synthesized, mainly to improve the solubility properties of chitosan (Progress in Polymer Science 26, 2001, 1921; Progress in Polymer Science 29, 2004, 887). Anionic water soluble chitosan derivatives are carboxyl acid derivatives (International Journal of Biological Macromolecules 14, 1992, 122; European Polymer Journal 39, 2003, 1629), phosphates (Carbohydrate Polymers 44, 2001, 1) and sulfates (Carbohydrate Research 302, 1997, 7). Other water soluble chitosan derivatives are poly(ethylene glycolated) derivatives (Carbohydrate Polymers 36, 1998, 49).
Important water soluble chitosan derivatives are derivatives with a quaternary ammonium moiety. These derivatives have two major advantages over the parent chitosan: (1) they are water-soluble on a wide pH-range including neutral and basic conditions, and (2) they have a permanent positive charge on the polymer backbone. The polycationic nature is commonly regarded to be responsible for the unique properties and activity of chitosan. Quaternary chitosan derivatives can be prepared either by quaternizing the amino group already present in the polymer or by adding a quaternary ammonium moiety or moieties. Synthesis of (N,N,N)-trimethylchitosan has been widely studied and reported (Carbohydrate Polymers 5, 1985, 297; International Journal of Biological Macromolecules 8, 1986, 105; Carbohydrate Polymers 24, 1994, 209; Carbohydrate Polymers 36, 1998, 157; Drug Development and Industrial Pharmacy 27, 2001, 373). The pharmaceutical properties of (N,N,N)-trimethylchitosan have been widely studied (e.g., European Journal of Pharmaceutics and Biopharmaceutics 58, 2004, 225; Biomaterials 23, 2002, 153; Carbohydrate Research 333, 2001, 1). However, well-defined uniform chitosan derivative structures cannot be obtained by direct methylation if the hydroxyl groups are not protected. Hydroxyl groups in the polymer, i.e., the primary hydroxyl at position 6 and the secondary hydroxyl at position 3, are also methylated. High degrees of quaternization cannot be obtained without the total O-methylation of the polysaccharide (Carbohydrate Polymers 36, 1998, 157).
The amino group in chitosan has also been quaternized by first reductively alkylating it with aldehydes to form imines, followed by reduction to obtain N-alkyl derivatives. These alkyl derivatives have been further quaternized with alkyl iodides (Polymer Bulletin 38, 1997, 387; Carbohydrate Research 333, 2001, 1; European Polymer Journal 40, 2004, 1355). Uragami and co-workers have crosslinked (N,N,N)-trimethylchitosan with various crosslinking agents, e.g., with tetraethoxysilane (Biomacromolecules 5, 2004, 1567) and with diethylene glycoldiglycidylether (Macromolecular Chemistry and Physics 203, 2002, 1162). Murata et al. quaternized some of the amino groups in galactose derivative of chitosan (Carbohydrate Polymers 29, 1996, 69; Carbohydrate Polymers 32, 1997, 105). Ucheqbu et al. prepared a quaternary ammonium palmitoyl chitosan to obtain a polysoap for drug delivery (International Journal of Pharmaceutics 224, 185-199). However, all these share the problem with (N,N,N)-trimethylchitosan, i.e., uniform structures cannot be obtained due to methylation of the hydroxyl groups of the polymer during the synthetic procedure.
The quaternary ammonium moiety can be inserted into polymer structures via various spacers. N-[(2-hydroxy-3-trimethylammonium)propyl]chitosan chloride can be obtained by reacting chitosan with glycidyltrimethylammonium chloride (Biomaterials 24, 2003, 5015; Carbohydrate Research 339, 2004, 313; Coloration Technology 120, 2004, 108; Colloids and Surfaces A: Physicochemical Engineering Aspects 242, 2004, 1; Polymer Journal 32, 2000, 334; International Journal of Biological Macromolecules 34, 2004, 121-126). This N-[(2-hydroxy-3-trimethylammonium)propyl]chitosan has been studied for different applications such as in cosmetics (e.g., U.S. Pat. No. 4,772,690; U.S. Pat. No. 4,822,598; U.S. Pat. No. 4,976,952). This derivative with varying lengths of alkyl chains attached to quaternary nitrogen has also been described as an antimicrobial agent (U.S. Pat. No. 6,306,835) and as a cholesterol lowering agent (WO9206136).
Another example of quaternary chitosan derivatives is N-betainate chitosan (Macromolecules 37, 2004, 2784; S.T.P. Pharma Sciences 8, 1998, 291). Lee et al. prepared quaternized diaminoalkylchitosans to obtain chitosan derivatives having two quaternary moieties (Bioscience Biotechnology and Biochemistry 63, 1999, 833; Bioorganic & Medicinal Chemistry Letters 12, 2002, 2949). Chun-Ho et al. prepared and studied the antibacterial activity of (triethylaminoethyl)chitin (Polymers for Advanced Technologies 8, 1997, 319). Suzuki et al. prepared N-p-(N-methylpyridinio)methylated chitosan and N-4-[(3-trimethylammonio)propaxy]benzylated chitosan and studied the electric resistance of these materials (Polymer Journal 32, 2000, 334).
Other polysaccharides have also been modified by inserting a quaternary ammonium moiety, e.g., cellulose (Macromolecular Materials and Engineering 286, 2001, 267) and starch (International Journal of Biological Macromolecules 31, 2003, 123). There are several commercial producers of these water-soluble quaternary derivatives of starch and cellulose. Tsai et al. reported alkylation of starch with monoquaternary 4,4-diethyl-1-(chloroethyl)piperazinium chloride hydrochloride and with diquaternary 1-glycidyl-1,4,4-trimethylpiperazinium dichloride (U.S. Pat. No. 5,349,089). However, no physicochemical properties, e.g., aqueous solubility of these alkylated starch derivatives were reported.
We have earlier prepared non-quaternary N-methylpiperazine derivatives of chitosan, but these were only relatively more soluble in water than the parent chitosan (Biomacromolecules 6, 2005, 858). By preparing quaternary piperazine derivatives we can obtain derivatives that are highly water-soluble on a wide pH range. However, quaternary piperazine derivatives cannot be prepared directly from these non-quaternary chitosan derivatives. To obtain a quaternary nitrogen atom one needs harsh reaction conditions usually with a large excess of the alkylating reagent. It is impossible to obtain well-defined chitosan derivatives by alkylating the non-quaternary chitosan derivatives, since this approach would result in a heteropolymer with both diquaternary and monoquaternary piperazine moieties in monomers. Also the hydroxyl groups in chitosan would be alkylated. The alkylation of hydroxyl groups of chitosan have shown to decrease the aqueous solubility of chitosan, e.g., Sieval et al. reported that the quaternary chitosan derivatives with high degrees of O-methylation were insoluble in water, even with high degree of quaternization (Carbohydrate Polymers 36, 1998, 157). This also proves that quaternary chitosan derivatives, even with high degree of quaternization, are not necessary water-soluble.